Kawasaki syndrome ("KS" hereafter) is an acute multi system vasculitis of unknown etiology. The disease primarily affects infants and young children, i.e., aged sixteen or younger. See Kawasaki, Jpn. J. Allergol 16: 178-222 (1967); Rauch et al., Pediatr. Infect. Dis. 4: 702-702 (1985). While KS does occur worldwide, it is most prevalent in Japan and in children of Japanese ancestry. Primary clinical manifestations include prolonged fever, bilateral non-exudative conjuctivitis, induration and erythema of extremities, inflammation of lips and oropharynx, polymorphous skin rash, and cervical lymphadenopathy. These indications are used in a clinical diagnosis of KS.
In Japan and in the United States, KS has become one of the most common causes of acquired heart disease in children. Recent studies have shown that when gamma globulin is administered intravenously ("IVGG") during the acute phase of the disease, coronary artery lesions, which otherwise develop in 15-25% of patients, are significantly decreased. See Newburger et al., N. Engl. J. Med. 315: 341-6 (1986); Nagashima et al., J. Pediatr. 110: 710-2 (1987); Firisho et al., Lancet ii: 1055-57 (1984); Rowley et al., J. Pediatr. 113: 290-94 (1988); Newburger et al., N. Eng. J. Med. 324: 1633-39 (1991). Thus, in order to treat this disease effectively, as with all other vasculitic diseases, early recognition is essential.
KS is characterized by an acute stage, as well as a convalescent stage. The acute phase is characterized, inter alia, by marked immune activation. Investigators have demonstrated, for example, increased number of circulating and infiltrating T cells bearing the HLA-DR activation antigen and elevated serum soluble IL-2 receptor levels. These phenomena are indicative of T-cell activation. See Leung et al., J. Clin. Invest. 79: 468-472 (1987); Terai et al., Hum. Pathol. 21: 231-234 (1990); Lang et al., J. Pediatr. 116: 592-596 (1990). In addition, acute KS has been associated with increased production of IL-1B, TNF-.alpha., IL-6, IL-2, and IFN-.gamma.. See, e.g., Matsubara et al., Clin. Immunol. Immunopathol 56: 29-36 (1990); Maury et al., J. Lab. Clin. Med. 113: 651-54 (1989); Lang et al., J. Pediatr. 115: 939-43 (1989); Leung et al., Lancet ii: 1928-1302 (1989); Rowley et al., Ped. Inf. Dis. J. 7: 663-67 (1988); Ueno et al., Clin. Exp. Immunol 76: 337-342 (1989); Jordan et al., in Kawasaki, ed., The Third International Kawasaki Disease Symposium 1989: 144-46. The cytokines referred to supra are believed to play a significant role in the pathogenesis of vascular cell injury during acute KS, due to their proinflammatory and prothrombotic effect on endothelial cells. See Mantovani et al., Immunol. Today 10: 370-74 (1989). Vascular endothelium, in KS lesions, has been demonstrated to express cytokine inducible leukocyte adhesion molecules known to be involved in localization of inflammatory cells. See Leung, supra. Patients with acute KS have been found to have cytotoxic antibodies against IL-1B, TNF-.alpha. and IFN-.gamma. stimulated endothelial cells, but not unstimulated cells. See Leung et al., J. Clin. Invest. 77: 1428-35 (1986); Leung et al., J. Exp. Med. 164: 1958-72 (1986).
While epidemiologic studies directed toward identifying potential environmental toxins, and laboratory culturing of body fluids for known microbial agents have taken place, an etiological agent for KS has not been found. See Rauch et al., Ped. Infect. Dis. J. 6: 1016-21 (1987). Due to the acute, self-limited nature of the disease, geographic clustering of outbreaks, clinical symptoms of fever and eruptions which mimic conditions and diseases such as measles, roseola, and scarlet fever, as well as the unique susceptibility of young children, it has been suggested that humoral immunity to this organism develops early in life. KS is rarely seen over the age of 8 suggesting that there is an asymptomatic infection caused by a ubiquitous agent, followed by development of protective immunity in the general population.
The general observations on KS, suggest that this disease has some similarities with disorders characterized by response to a so-called "superantigen". The previously cited references show that various superantigens lead to expanded populations of V.beta. elements or TCRs ("T cell receptor molecules"). This evidence is also presented in, e.g., Choi et al., J. Exp. Med. 172: 981-84 (1990); Kappler et al., Science 24: 811-13 (1989); and Choi et al, Proc. Natl. Acad. Sci. 86: 8941-45 (1989). The disclosures of these three references are all incorporated by reference herein. The superantigens, including bacterial toxins, provoke marked activation of T cells and monocytes/macrophages. For example, staphylococcal enterotoxins and streptococcal erythrogenic toxins induce IL-1 and TNF-.alpha. from monocytes. Staphylococcal enterotoxin and SPE-mediated stimulation of monocytes is a consequence of binding and transducing a positive signal through MHC-II molecules on monocyte cell surfaces. In the presence of antigen presenting cells, superantigens stimulate T-cell proliferation by selective stimulation of T cells expressing particular V.beta. elements. For example, Staphylococcal TSST-1 stimulates T cells presenting V.beta.2. Choi et al., J. Exp. Med. 172: 981-4 (1990), have shown expansion of V.beta.2 cells in toxic shock syndrome. The similarities thus suggest at this time that vasculitic diseases especially KS, may involve the same phenomena as is involved in superantigen caused diseases and conditions, but, as noted supra, this is a theory rather than a known mechanism, as compared to the expansion of the V.beta. subtype, which is an empirical phenomenon.
Abe et al., Proc. Natl. Acad. Sci. USA 89: 4066-4070 (5/92), the disclosure of which is incorporated by reference in its entirety, describe experiments wherein the T cell repertoire of patients with KS were studied. It was found that the variable regions V.beta.2 and, to a lesser extent, V.beta.8, were expanded within these patients, relative to controls and to other variable regions. The paper reiterates the discussion supra, i.e., that the cause of KS is unknown. The paper speculates that streptococcal exotoxins or homologous exotoxins may be involved in the pathogenesis of acute KS.
The parent of the subject application disclosed that a diagnosis of Kawasaki Syndrome can be made by assaying for Streptococcal bacteria, and its associated antigens, or by assaying for Staphyloccal bacteria which produce toxic shock syndrome toxin "TSST-1". More precisely, a strain of S. aureus which differs from all other previously observed strains has been identified. The implicated strain is a white color in appearance. It was observed that the cultures appeared benign, but were involved in pathological conditions. The observations suggested that other undiagnosed disorders in addition to KS may be associated with pathogenic bacteria which appear to be normal.
The present invention involves the isolation of a nucleic acid molecule which codes for a toxin associated with Kawasaki Syndrome. The toxin has a particular amino acid sequence, which is set forth, in SEQ ID NO: 1. The invention involves isolated nucleic acid molecules which code for proteins having the nucleotide sequence, set forth in SEQ ID NO: 2. Also disclosed are such nucleic acid molecules. Other features of the invention include vectors which comprise the nucleic acid molecules of the invention operably linked to promoters, as well as cell lines, such as prokaryotic (e.g., E. coli) and eukaryotic (e.g., CHO and COS) cells transfected with the nucleic acid molecule or with the promoters of the invention. The identification of the molecule and its coding sequence also enables the artisan to identify organisms implicated in Kawasaki Syndrome, by the use of specific probes.